Preliminary evaluation of pharmacological properties of some xanthone derivatives

Article abstract of DOI:10.1016/j.bmc.2008.12.031

A series of xanthone derivatives were synthesized and examined for electrocardiographic, antiarrhythmic, hypotensive and anticonvulsant activities as well as for α₁- and β₁-adrenergic binding affinities. Among the investigated compounds, some of them exhibited significant antiarrhythmic and/or hypotensive activity. The data obtained via receptor binding assay are in agreement with pharmacological results and could explain antiarrhythmic and/or hypotensive activity of the newly synthesized structures.

Full text of DOI:10.1016/j.bmc.2008.12.031

Bioorganic & Medicinal Chemistry 17 (2009) 1345–1352
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry
journal homepage: www.elsevier.com/locate/bmc
Preliminary evaluation of pharmacological properties of some
xanthone derivatives
Henryk Marona , Natalia Szkaradek , Anna Rapacz , Barbara Filipek c,d, Małgorzata Dybała e,
a,*
a,b
c
e
b
f
Agata Siwek , Marek Cegła , Edward Szneler
Department of Technology and Biotechnology of Drugs, Faculty of Pharmacy, Jagiellonian University Medical College, 9 Medyczna Str., 30-688 Cracow, Poland
Department of Organic Chemistry, Faculty of Pharmacy, Jagiellonian University Medical College, Cracow, Poland
Department of Pharmacodynamics, Faculty of Pharmacy, Jagiellonian University Medical College, Cracow, Poland
Laboratory of Pharmacologicall Screening, Faculty of Pharmacy, Jagiellonian University Medical College, Cracow, Poland
a
b
c
d
e
f
Department of Cytobiology and Histochemistry, Laboratory of Pharmacobiology, Faculty of Pharmacy, Jagiellonian University Medical College, Cracow, Poland
Faculty of Chemistry, Jagiellonian University, Cracow, Poland
a r t i c l e i n f o
a b s t r a c t
Article history:
Received 30 July 2008
Revised 3 December 2008
Accepted 8 December 2008
Available online 24 December 2008
A series of xanthone derivatives were synthesized and examined for electrocardiographic, antiarrhyth-
mic, hypotensive and anticonvulsant activities as well as for - and b -adrenergic binding affinities.
Among the investigated compounds, some of them exhibited significant antiarrhythmic and/or hypoten-
sive activity. The data obtained via receptor binding assay are in agreement with pharmacological results
and could explain antiarrhythmic and/or hypotensive activity of the newly synthesized structures.
Ó 2008 Elsevier Ltd. All rights reserved.
a
1
1
Keywords:
Xanthones
Antiarrhythmic, hypotensive and
anticonvulsive activities
Radioligand-binding assay
1
. Introduction
tain an 1-aroxy-3-alkylamino-2-propanol group in their structure
(
propranolol, atenolol, metoprolol, acebutolol). Furthermore 1-ar-
oxy-3-alkylamino-2-propanol group is structural element of some
nonselective, third generation b-blockers with additional -adre-
Cardiovascular diseases are caused by disorders of the heart and
blood vessels, and include coronary heart disease, cerebrovascular
disease, hypertension, peripheral artery disease, rheumatic heart
disease, congenital heart disease and heart failure. Although, devel-
opment of modern, effective therapies, cardiovascular diseases re-
mains the number one cause of death, representing 30% of all
global deaths. Hypertension and arrhythmia are the major factors
for cardiovascular mortality, which reach nearly three quarter of
a
nolytic properties (carvedilol) revealing beneficial effect on lower-
ing blood pressure. Taking into account these facts, publications
5
concerning cardiovascular properties of aminoalcanolic derivatives
6
–8
9–11
of xanthone
and our own experience,
herein we reported the
results of in vivo evaluating of potential antiarrhythmic and hypo-
tensive activity of selected xanthone derivatives.
1
,2
them. In the genesis of cardiac dysfunctions, an important role
plays activation of sympathetic nervous system and from here
adrenergic receptors antagonists have been widely accepted in
It is known from the literature that the anticonvulsant activity
has been demonstrated for compounds without any structural sim-
ilarity to classic antiepileptic drugs (valproic acid, carbamazepine,
lamotrigine, etc.). Significant example can be propranolol having
an established indication in a variety of cardiovascular diseases
and interestingly, potently preventing maximal electroshock
(MES) seizures in rodents and raising the after discharge threshold
3
,4
treatment. On the other hand, an ideal antiarrhythmic or hypo-
tensive medicament does not exist. Treatment is associated with
side-effects, such as tiredness, changes in mood, sleep distur-
bances, dry mouth, blurry vision or impotence. Thus, delicate bal-
ance between drug efficacy and unexpected adverse side effects is
essential. It results in importance of searching for the new agents
improving heart function with minimal side effects.
1
2
in rats. Among these compounds there are also aminoalcanols,
for which a high anticonvulsant activity has been described for
example
2-(3N-(tert-butylamino)-2-hydroxypropoxy)-9H-xan-
It is known that most classic b-blockers, classified as second
class of antiarrhythmic drugs according to Vaughan Williams, con-
then-9-one revealing protection index (PI) in maximal electro-
shock-induced seizures (mice, i.p.) ranging 4.5, which is
1
0
comparable with that for Carbamazepine (PI = 4.9). These knowl-
edge prompted us to carry out additional tests evaluating neuro-
toxicity and potential antiepileptic properties of the newly
*
Corresponding author. Tel./fax: +48 12 657 04 88.
E-mail address: hen.mar@interia.pl (H. Marona).
0
968-0896/$ - see front matter Ó 2008 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmc.2008.12.031

1
346
H. Marona et al. / Bioorg. Med. Chem. 17 (2009) 1345–1352
synthesized compounds. Preliminary pharmacological and neuro-
toxicological assays have been provided by the ADD (Anticonvul-
sant Drug Development) program, Epilepsy Branch, National
Institute of Neurological Disorders and Stroke, National Institute
of Health (Rockville, USA).
Table 1
Chemical structures of the tested compounds
O
8
1
7
2
3
R2
·
HCl
6
O
4
R1
5
2
2
. Results
Compound
R
1
R
2
2
.1. The influence on the normal electrocardiogram
The effect on ECG intervals and the heart rate was determined
OH
OH
OH
OH
H
N
1
H
H
H
O
O
O
O
CH2
for all compounds at the doses of 5 or 10 mg/kg body weight
Table 1). Electrographic experiments showed that compounds 4,
, 6, 8 significantly decreased the number of cardiac beats per min-
(
5
H
N
2
3
3
4
CH2
ute—4 by 13%, 5 by 20%, 6 by 15%, and 8 by 21%, besides com-
pounds 4, 5 and 6 prolonged the PQ interval, especially in the
first minute of the test (Figs. 1 and 2). Compounds 3 and 7 signif-
icantly decreased the number of cardiac beats per minute (24% and
H
N
CH2
3
0%) and prolonged the QT interval (17% and 13%) during all the
time of observation (Fig. 3). The compound 1 diminished the heart
rate and prolonged the PQ interval only in the first minute of the
test (Fig. 3). The compound 2 did not significantly affect the normal
ECG, only slightly prolonged the QT interval.
H
N
CH OH
4
5
6
7
8
4
4
4
4
4
2
H
CH3
CH OH
2
H
N
2
.2. Antiarrhythmic activity
O
O
H
CH2
Prophylactic antiarrhythmic activity of the tested compounds
was evaluated using anesthetized rats in the adrenaline-induced
model of arrhythmia. Rapid intravenous injection of adrenaline at
a dose of 20
lg/kg caused reflex bradycardia (100%), atrioventricu-
lar disturbances, extrasystoles (88%), which led to the death of
approximately 76% of animals of the control group within 15 min
of the observation.
The tested compounds, injected intravenously 15 min before
adrenaline, diminished the occurrence of heart-rhythm
disturbances.
Among the studied compounds, the strongest prophylactic anti-
arrhythmicactivitywasshownbycompounds4and7.TheirED50 val-
ues (a dose producing a 50% inhibition of premature ventricular
beats) are presented in Table 2. These data show that compound 4
was the most active in this test. This compound as well as 7 also pre-
vented the appearance of other adrenaline-induced arrhythmia
symptoms(bradycardia, blocks,bigeminy)andsignificantlyreduced
mortality. The compound 7 at the lowest dose in this test (1.25 mg/
kg)stillreducedmortalityby60%incomparisontothecontrolgroup.
H
N
CH
3
H
CH OH
2
CH3
OH
OH
H
N
CH3
O
O
Cl
Cl
CH OH
2
CH3
H
N
CH OH
2
CH3
CH OH
2
both systolic (22–10%) and diastolic (25–11%) blood pressure,
which persisted for 60 min.The results for these compounds are
presented in Figures 4 and 5. Compounds 2 and 6 at a dose of
1
0 mg/kg significantly influenced blood pressure only in 60 min
2
.3. Influence on blood pressure
of the test. Compound 5 (at a dose of 10 mg/kg) initially reduced
systolic and diastolic pressure, but the pressure returned to the
baseline value within next couple of minutes after administration.
Compounds 4 and 8, at a dose of 2.5 mg/kg, had no effect in this
test (data not shown).
The investigated compounds were tested for hypotensive activ-
ity after intravenous administration at the doses of 10 mg/kg (1, 2,
, 5, 6, 7) or 2.5 mg/kg (4 and 8). For active compounds, these doses
3
were reduced until the hypotensive activity disappeared.
Intravenous injections of compound 7 at doses of 10–1.25 mg/
kg significantly reduced the systolic and diastolic blood pressure
in normotensive rats and that hypotensive effect lasted throughout
the observation. At a lower dose (0.625 mg/kg), the compound 7
significantly reduced the blood pressure only in 60 min of the
observation (data not shown). Compound 7 reduced systolic blood
pressure by 27–17% and diastolic blood pressure by 33–19% at the
lowest dose used in this test (1.25 mg/kg).
2.4. Radioligand-binding assay
3
Compounds 3, 4, 7 and 8 displaced [ H]CGP-12177 from cortical
binding sites in low concentration range (K
compounds moderately inhibited [ H]CGP-12177 binding with
i
= 3.9–83.8 nM). Other
3
3
l
M-range. All compounds inhibited [ H]prazosin binding to corti-
cal -adrenoceptors with M-range. The results are summarized
in Table 2.
a
1
l
Compound 3 reduced the systolic blood pressure by 16–9% and
diastolic blood pressure by 18–11% at a dose of 5 mg/kg. At a lower
dose (2.5 mg/kg), the compound 3 did not have any significant
influence on blood pressure (data not shown). Compound 1 re-
vealed hypotensive activity only at a dose of 10 mg/kg; it reduced
2.5. Anticonvulsant and neurotoxicity assay
Synthesized compounds were evaluated for anticonvulsant
activity in the maximal electroshock (MES)- and subcutaneous

H. Marona et al. / Bioorg. Med. Chem. 17 (2009) 1345–1352
1347
Table 2
5
6
4
8
(10mg/kg)
(10mg/kg)
(5mg/kg)
(5mg/kg)
400
350
300
250
200
3
3
K
i
values for the inhibition of the binding of [ H]prazosin and [ H]CGP-12177 to
1
a -
and b -adrenoceptors and the prophylactic antiarrhythmic activity in adrenaline
1
induced arrhythmia in anesthetized (thiopental 75 mg/kg i.p.) male Wistar rats (route
of administration—i.v.)
*
***
**
**
Compound
Ki ± SEMa
M)
1
-Adrenoceptors b -Adrenoceptors
(l
ED50b (mg/kg)
*
**
****
*
*
a
1
*
*
**
****
***
***
****
1
2
3
4
5
6
7
8
4.9 ± 1.1
8.1 ± 1.6
7.3 ± 0.5
49.8 ± 5.5
4.6 ± 0.5
8.7 ± 0.2
5.6 ± 0.6
20.6 ± 2.1
—
30.9 ± 0.7
44.4 ± 4.2
0.084 ± 0.010
0.004 ± 0.0007
7.7 ± 0.3
—
—
—
*
***
****
****
1.753 (0.77–4.01)
***
—
—
2.6 ± 0.3
0
1
5
10
15
0.011 ± 0.003
0.047 ± 0.004
9.7 ± 0.8
2.46 (1.05–5.73)
—
1.05 (0.64–1.73)
Time (min)
Propranolol
Figure 1. Effect of compounds 4, 5, 6 and 8 on electrocardiogram (cardiac beats per
a,b
K
i
± SEM values were derived from 2 to 3 experiments in duplicates.
minute) in anaesthetized rats after iv administration. Statistical analyses were
b
Each value was obtained from three experimental groups. Each group consisted
***
****
performed using a one-way ANOVA test: p < 0.01,
p < 0.001.
of six animals. The ED50 values and their confidence limits were calculated
according to the method of Litchfield and Wilcoxon.
44
42
40
38
36
*
**
160
7
(1.25mg/kg)
*
***
150
3 (5mg/kg)
1
(10mg/kg)
*
*
*
140
130
120
110
100
***
***
****
***
*
***
*
****
***
****
***
*
**
**
*
**
****
****
*
5 (10mg/kg)
*
6
4
(10mg/kg)
(5mg/kg)
*
**
*
**
0
1
5
10
15
**
***
90
***
Time (min)
Figure 2. Effect of compounds 4, 5 and 6 on electrocardiogram (PQ intervals per
80
0
1
5
10
20
30
40
60
minute) in anaesthetized rats after iv administration. Statistical analyses were
*
**
***
****
performed using a one-way ANOVA test: p < 0.05, p < 0.02, p < 0.01, p < 0.001.
Time (min)
Figure 4. The influence of compounds 1, 3 and 7 on the systolic blood pressure in
anaesthetized rats after iv administration. Statistical analyses were performed
4
4
3
3
2
2
50
00
50
00
50
00
7 (10mg/kg)
*
**
***
****
using a one-way ANOVA test: p < 0.05, p < 0.02, p < 0.01,
p < 0.001.
3
1
(10mg/kg)
(10mg/kg)
140
130
120
*
*
**
****
***
*
***
****
****
****
*
****
***
110
100
****
****
****
****
15
*
***
****
**
***
*
***
*
**
0
1
5
10
9
0
0
Time (min)
*
**
8
*
Figure 3. Effect of compounds 1, 3 and 7 on electrocardiogram (cardiac beats per
minute) in anaesthetized rats after iv administration. Statistical analyses were
performed using a one-way ANOVA test: p < 0.01,
7 (1.25mg/kg)
3 (5mg/kg)
*
**
***
****
p < 0.001.
70
1
(10mg/kg)
60
0
1
5
10
20
30
40
60
pentylenetetrazole (ScMet)-induced seizures tests and for neuro-
toxicity (TOX) in the rotorod test in mice (i.p.). Among the tested
compounds only 1 and 2 revealed some anti-MES activity (after
Time (min)
Figure 5. The influence of compounds 1, 3 and 7 on the diastolic blood pressure in
0
3
.5 and 4 h of i.p. administration, mice) in the doses of 100 and
00 mg/kg (see Table 3). None of the tested compounds displayed
anaesthetized rats after iv administration. Statistical analyses were performed
*
**
***
****
using a one-way ANOVA test: p < 0.05, p < 0.02, p < 0.01,
p < 0.001.
anti-ScMet protection (data not published). On the other hand,
most of the examined xanthone derivatives did not show neuro-
toxicity in the dose of 30 mg/kg (compounds 1, 2, 4, 5, 6, 8) as well
as in the dose of 100 mg/kg (compounds 1, 2, 4, 6, 8). Except 8, all
of the evaluated compounds revealed diverse (from 25% to 100%)

1
348
H. Marona et al. / Bioorg. Med. Chem. 17 (2009) 1345–1352
range of neurotoxicity after 0.5 h of administration in the dose of
cated that compounds with no hydroxylic groups were inactive
neither in reducing blood pressure nor in cardioprotection in the
adrenaline model of arrhythmia (data not published). On the other
hand, the most potent in the adrenaline model of arrhythmia were
compounds 4 and 7, with the ED50 value 1.75 mg/kg for compound
3
00 mg/kg.
3
. Discussion
4
and 2.46 mg/kg for compound 7. Propranolol, which was used in
The aim of this study was to evaluate cardiovascular activity of
eight new xanthone derivatives. This work includes synthesis and
pharmacological studies of these compounds. It examines both car-
this test as a reference compound had ED50 value 1.05 mg/kg. From
the radioligand-binding assay it appeared that compounds 4 and 7
had high affinity to b -adrenergic receptors in rat cerebral cortex (4
1
and 11 nM, respectively, see Table 3). Based on our results, we sug-
gest that the antiarrhythmic effects of the tested compounds are
diovascular activity: anti-arrhythmic, hypotensive,
1 1
a - and b -
adrenergic blocking activities, effect on the normal electrocardio-
gram and influence on central nervous system.
1
related to the b -adrenoceptor blockade in heart tissue.
Searching for new xanthone derivatives with potential biologi-
cal properties has remained in the area of our interest for the last
several years and has been documented by a number of publica-
tions. Among the evaluated compounds there were structures
However, compound 8 in comparison with 7 was less active.
Analogously compound 4 was less active than its unhydroxylated
analogue: 4-(3N-(2-amino-2-methyl-1-hydroxypropyl)-2-hydro-
13–15
16
1
xypropoxy)-9H-xanthen-9-one hydrochloride revealing b -affinity
revealing for example anticonvulsant
, antitumour, antitu-
1
8
berculotic, aniarrhythmic and/or hypotensive9–11 activity. Thus
our own experience and former articles which treated xanthone
derivatives possessing cardiovascular activity6 prompted us to
synthesize a novel series of xanthone derivatives with aminoalcan-
olic moieties—well known structural element of b-blockers. Addi-
tionally we synthesized and evaluated their two aminic
analogues. Results obtained via antiarrhythmic, hypotensive and
binding affinity assays confirmed that aminoalcanolic moiety play
an important role in the structure of cardiovascular drugs. For that
reason, compounds 5 and 6 were less active than their alcanolic
derivatives. Presence of one hydroxylic group seems to be essential
also in the amine molecule. Fact that many of b-blockers possess
iso-propylamine or tert-butylamine moiety in their structure
encouraged us to synthesize their xanthonic analogues and xanth-
onic analogues with additional hydroxylic groups. Evaluation of
the obtained substances towards cardiovascular properties indi-
17
amounted to 2.6 nM. These two facts suggested that presence
of one hydroxylic group is favourable to potential antiarrhythmic
properties of the tested compound. Nevertheless, presence of two
hydroxylic group and in consequence lower lipophilicity caused
decreasing of efficacy.
Concerning influence on blood pressure, the most potent seem
to be compounds 3 and 7, containing allylamine moiety, new, po-
tential hypotensive farmacophore. Compound 7 significantly re-
duced the systolic and diastolic blood pressure at the dose
–8
1
.25 mg/kg. Compound 3 decreased blood pressure all the time
during the observation but at higher doses. In binding studies com-
pounds 3 and 7 had additionally moderately high affinity for
adrenoceptors in binding studies. The hypotensive effect is proba-
bly the result of b -adrenoceptor blockade in heart and -adreno-
ceptor blockade in arteries.
a
1
1
a
1
Among all of the tested compounds with allylamine group,
place of substitution played a key role. Curiously enough, the most
required was position 4 in the xanthone rings, while according to
the former publication treating xanthonolol as a potential hypo-
tensive agent, the most suitable was substitution in the 3 position.
1
9
Table 3
Anticonvulsant screening project (ASP): phase I-test results in mice (i.p.
administration)
Compound
Dose (mg/kg)
Activity MESa time (h)
TOXb time (h)
4. Conclusion
0.5
4
0.5
4
The results confirmed that among the investigated compounds,
some of them exhibited significant b -adrenolitic, antiarrhythmic
1
1
30
0/1
2/3
1/1
0/1
0/3
1/1
0/4
0/8
4/4
0/2
0/4
1/2
1
3
00
00
and/or hypotensive activities. The strongest anti-arrhythmic activ-
ity in the adrenaline-induced model of arrhythmia revealed com-
pounds 4 and 7. The most potent hypotensive activity in our
research has the compound 7. The results obtained from radioli-
gand binding assays showed that tested compounds had affinity
2
3
4
5
6
7
8
a
30
0/1
1/3
1/1
0/1
0/3
1/1
0/4
0/8
4/4
0/2
0/4
0/2
1
3
00
00
30
0/1
0/3
0/1
0/1
0/3
0/1
1/4
1/8
2/4
0/2
0/4
0/2
1
3
00
00
to adrenoceptors. For compounds 3, 4, 7 and 8 the K
in nM range towards b adrenoceptors. The obtained data con-
cluded, that binding studies are in agreement with our pharmaco-
logical results and we put forward hypothesis that the
i
value was
1
30
0/1
0/3
0/1
0/1
0/3
0/1
0/4
0/8
1/4
0/2
0/4
0/2
1
3
00
00
a
antiarrhythmic and hypotensive activity are related to adrenolitic
properities of examined compounds.
30
0/1
0/2
—
0/1
0/2
—
0/4
5/8
4/4
0/2
0/3
—
1
3
00
00
30
0/1
0/3
0/1
0/1
0/3
0/1
0/4
0/8
1/4
0/2
0/4
0/2
5. Experimental
1
3
00
00
5
.1. Chemistry
30
0/1
0/3
0/1
0/1
0/3
0/1
1/4
5/8
4/4
0/2
1/4
2/2
1
3
00
00
Reagents: epichlorohydrin, allylamine, 2-amino-2-methyl-1-
propanol, 2-amino-2-methyl-1,3-propanediol were purchased from
Sigma–Aldrich Chemie (Steinheim, Germany), wherease reagents:
30
0/1
0/3
0/1
0/1
0/3
0/1
0/4
0/8
0/4
—
0/5
—
1
3
00
00
3
-chloro-1-propanol, phosphorus tribromide were provided by Lan-
caster Synthesis (Frankfurt am Main, Germany). Solvents were com-
mercially available materials of reagent grade. Melting points (mp)
are uncorrected and were determined using a Büchi SMP-20 appa-
ratus (Büchi Labortechnik, Flawil, Switzerland). The infrared spectra
Maximal electroshock test (number of animals protected/number of animals
tested).
Rotorod toxicity (number of animals exhibiting toxicity/number of animals
tested).
b

H. Marona et al. / Bioorg. Med. Chem. 17 (2009) 1345–1352
1349
were recorded on potassium bromide pellets using a Jasco FT/IR 410
spectrometer (Jasco Inc., Easton, MD, USA). The 1H NMR for com-
pounds 3, 5, 6, 7 were recorded on a Bruker AMX spectrometer
(DMSO-d
3.75 (t, J = 6.4, 2H, –CH
7.21–8.37 (m, 7H, Ar–H). R
6
, 300 MHz) d (ppm): 2.48 (m, 2H, –CH
–Br), 4.32 (t, J = 5.9 Hz, 2H, –O–CH
= 0.93 (toluene/acetone (5:3)).
2
–CH
2
–CH
2
–),
–),
2
2
f
(
Brucker, Karlsruhe, Germany) with 500.13 MHz, using signal from
DMSO in DMSO-d and TMS in CDCl as an internal standard,
whereas H NMR spectra for the other compounds were obtained
in CDCl or DMSO-d with a Varian Mercury-VX 300 NMR spectrom-
5.2.2. Synthesis of the tested compounds
6
3
1
Racemic compounds 1–8 (see Table 1) were obtained as pre-
sented in Scheme 1 by amination of respective parent compound
with appropriate amines in n-propanol (for 1–4, 7, 8) or toluene in
3
6
eter (Varian Inc., Palo Alto, CA, USA) with TMS or DMSO, respec-
tively, as an internal standard. The results are presented in the
following format: chemical shift d (ppm), multiplicity, J values in
Hertz (Hz), number of protons, proton’s position. Multiplicities were
shown as the abbreviations: s (singlet), br s (broad singlet), br b
a presence of K
2
CO
3
(for 5, 6), according to the earlier published pro-
9
,13
cedures.
Resulted bases were converted into hydrochloride salts
in propanol/acetone (4:1) with an excess of ethanol saturated with
HCl. The crude products were recrystallized from acetone/ethanol
(1:3).
(
broad bond), d (doublet), dd (doublet of doublets), td (triplet of
doublets), t (triplet), dt (doublet of triplets), tt (triplet of triplets),
qu (quintet), m (multiplet). Elemental analyses were performed
on a Elementar Vario EL III (Elementar Analysansysteme, Hanau,
Germany). The purity of obtained compounds was confirmed by
the thin-layer chromatography (TLC), carried out on precoated
plates (Silica Gel, 60 F-254, Merck, Darmstadt, Germany) using the
solvent systems mentioned below. The obtained corresponding
spots were visualized under UV light.
5.2.2.1. 2-(3-(Allylamino)-2-hydroxypropoxy)-9H-xanthen-9-one
hydrochloride (1) was obtained as white solid (yield 70%), mp
206–207 °C. Anal. Calcd for C19
3.87. Found: C, 63.06; H, 5.72; N, 3.75. IR (KBr)
2932, 2788, 1619, 1467, 1215, 1037, 764. Base of compound 1:
4
H20NO Cl: C, 63.07; H, 5.57; N,
ꢀ
1
v
(cm ): 3384,
1
H NMR (CDCl
2.93 (m, 2H, –CH(OH)–CH
CH@CH ), 4.05–4.17 (m, 4H, Ar–O–CH
H, –CH@CH ), 5.85 (m, 1H, –CH@CH ), 7.35–8.36 (m, 7H, Ar–H).
= 0.28 (methanol/ethyl acetate (1:1)).
.2.2.2. 3-(3-(Allylamino)-2-hydroxypropoxy)-9H-xanthen-9-one
hydrochloride (2) was obtained as white solid (yield 64%), mp
184–186 °C. Anal. Calcd for C19 Cl: C, 63.07; H, 5.57; N,
3.87. Found: C, 63.04; H, 5.53; N, 3.75. IR (KBr)
3
, 300 MHz) d (ppm): 1.72 (br s, 1H, –NH–), 2.79–
–NH–), 3.26–3.38 (m, 2H, –NH–CH
–CH(OH)–), 5.11–5.25 (m,
2
2
–
2
2
2
R
2
2
5
5
.2. Chemical syntheses
f
5
.2.1. Synthesis of the starting materials
The synthetic route used to synthesize starting materials is out-
H20NO
4
ꢀ
1
lined in Scheme 1. The detailed description of the method and
physico-chemical properties of 2-((Oxiran-2-yl)methoxy)-9H-xan-
then-9-one, 3-((Oxiran-2-yl)methoxy)-9H-xanthen-9-one and 4-
v (cm ): 3266,
1
2929, 1622, 1467, 1283, 1166, 1106, 759. H NMR (DMSO-d
300 MHz) d (ppm): 2.94–3.15 (m, 2H, –CH(OH)–CH
3.61 (d, J = 6.4 Hz, 2H, –NH
Ar–O–CH
2H, –CH@CH
6
,
þ
2
–NH
–),
2
þ
(
(Oxiran-2-yl)methoxy)-9H-xanthen-9-one were described else-
2
–CH
2
–CH@CH
2
), 4.13–4.27 (m, 3H,
), 5.86–5.98 (m,
, –OH), 7.05–8.18 (m, 7H, Ar–H), 9.02 (br s, 1H, –
9
–11
where.
3-Chloro-5-((Oxiran-2-yl)methoxy)-9H-xanthen-9-one
2 2
–CH(OH)-), 5.38–5.51 (m, 2H, –CH@CH
was obtained analogously from 3-chloro-5-hydroxy-9H-xanthen-
-one. The crude products were recrystallized from ethanol. 4-(3-
2
+
+
9
f
NHH –), 9.19 (br s, 1H, –NHH –). R = 0.28 (methanol/ethyl acetate
bromopropoxy)-9H-xanthen-9-one was prepared from 4-hydro-
xy-9H-xanthen-9-one according to the previously described proce-
(1:1)).
5.2.2.3. 4-(3-(Allylamino)-2-hydroxypropoxy)-9H-xanthen-9-one
hydrochloride (3) was obtained as white solid (yield 65%), mp
1
1
dure using 3-chloro-1-propanol instead of 2-chloroethanol (see
Scheme 1). The crude products were recrystallized from n-hex-
ane/toluene (1:4).
163–165 °C. Anal. Calcd for C19
3.87. Found: C, 62.58; H, 5.68; N, 3.80. IR (KBr)
2942, 1651, 1594, 1467, 1278, 1082, 756. H NMR (DMSO-d
4
H20NO Cl: C, 63.07; H, 5.57; N,
ꢀ1
v
(cm ): 3357,
1
5
.2.1.1. 3-Chloro-5-hydroxy-9H-xanthen-9-one was obtained as
white solid (yield 70%), mp 174–176 °C. Anal. Calcd for C13 Cl:
(cm ): 3229,
6
,
H
7
O
3
500.13 MHz) d (ppm): 3.07 (dd, J = 8.7 Hz, J = 12.5 Hz, 1H, –CH–
ꢀ1
þ
C, 63.30; H, 2.86. Found: C, 63.34; H, 3.03. IR (KBr)
647, 1600, 1496, 1432, 1252, 1202, 1071, 934, 831. H NMR
DMSO-d , 300 MHz) d (ppm): 5.76 (s, 1H, –OH), 7.24–8.32 (m,
H, Ar–H). R = 0.91 (methanol/ethyl acetate (1:1)).
.2.1.2. 3-Chloro-5-((oxiran-2-yl)methoxy)-9H-xanthen-9-one was
obtained as white solid (yield 65%), mp 183–185 °C. Anal. Calcd
for C16 Cl: C, 63.48; H, 3.66. Found: C, 63.05; H, 4,01. IR (KBr)
(cm ): 3437, 3069, 1602, 1492, 1332, 1274, 1220, 1070, 928,
v
CHH–NH
2
–), 3.22 (dd, J = 3.5 Hz, J = 12.5 Hz, 1H, –CH–CHH–
1
þ
þ
1
NH
CH
2
–), 3.64 (ddd, J = 0.9 Hz, J = 1.2 Hz, J = 6.6 Hz, 2H, –NH
-), 4.22 (dd, J = 5.6 Hz, J = 10.2 Hz, 1H, Ar–O–CHH–), 4.26 (dd,
2
–
(
6
2
6
f
J = 4.9 Hz, J = 10.2 Hz, 1H, Ar–O–CHH–), 4.30–4.41 (m, 1H, –CH–),
5.40 (ddd, J = 0.9 Hz, J = 1.5 Hz, J = 10.2 Hz, 1H, @CHH), 5.49 (ddt,
J = 1.2 Hz, J = 1.5 Hz, J = 17.2 Hz, 1H, @CHH), 5.97 (ddt, J = 6.6 Hz,
5
H
O
11 4
2 2
J = 10.2 Hz, J = 17.2 Hz, 1H, –CH CH@CH ), 6.03 (br s, 1H, –OH),
ꢀ
1
v
7
7.40 (dd, J = 8.0 Hz, J = 8.0 Hz, 1H, 2Ar–H), 7.51 (ddd, J = 1.1 Hz,
J = 7.1 Hz, J = 8.0 Hz, 1H, 7Ar–H), 7.59 (dd, J = 1.5 Hz, J = 8.0 Hz,
1H, 3Ar–H), 7.71 (dd, J = 1.1 Hz, J = 8.7 Hz, 1H, 5Ar–H), 7.77 (dd,
J = 1.5 Hz, J = 8.0 Hz, 1H, 1Ar–H), 7.92 (ddd, J = 1.8 Hz, J = 7.1 Hz,
J = 8.7 Hz, 1H, 6Ar–H), 8.21 (dd, J = 1.8 Hz, J = 8.0 Hz, 1H, 8Ar–H),
1
6
53. H NMR (DMSO-d , 300 MHz) d (ppm): 2.87 (dd, J = 2.7 Hz,
J = 8.7 Hz, 1H, –CHH–), 2.99 (dd, J = 4.1 Hz, J = 4.9 Hz, 1H, –CHH–),
.47–3.52 (m, 1H, –CH@), 4.13 (dd, J = 5.9 Hz, J = 11.3 Hz, 1H,
Ar–O–CHH–CH@), 4.47 (dd, J = 2.8 Hz, J = 11.3 Hz, 1H, Ar–O–
3
þ
13
CHH–CH@), 7.26–8.28 (m, 6H, Ar–H). R
F
= 0.88 (methanol/ethyl
8.89 (br s, 2H, –NH
(ppm): 48.81 (R–CH
CHOH–R), 71.34 (Ar–O–CH
(C-5), 121.03 (C-8a), 122.16 (C-8b), 122.89 (@CH
124.69 (C-7), 126.09 (C-8), 129.27 (@CH–), 135.69 (C-6), 146.17
2
–). C NMR (DMSO-d
–NH–), 49.31 (–NH–CH –CH@), 64.92 (R–
-), 117.26 (C-1), 118.30 (C-3), 118.49
), 124.11 (C-2),
6
, 500.13 MHz) d
acetate (1:1)).
2
2
5
.2.1.3. 4-(3-Hydroxypropoxy)-9H-xanthen-9-one was obtained
as white solid (yield 65%), mp 136–137 °C. Anal. Calcd for
: C, 71.10; H, 5.22. Found: C, 71.02; H, 5.23. IR (KBr)
cm ): 3365, 2935, 1668, 1467, 1337, 1274, 1256, 1226, 1067,
2
2
C
(
16
H
14
O
4
v
ꢀ
1
f
(C-4a), 147.55 (C-4), 155.41 (C-4ba), 176.10 (C@O). R = 0.29
(methanol/ethyl acetate (1:1)).
1
9
2
–
40, 751. H NMR (DMSO-d
H, –CH –CH –CH –), 2.55 (t, J = 5.6, 1H, –OH), 3.97–4.03 (m, 2H,
CH –OH), 4.34 (t, J = 5.9 Hz, 2H, –O–CH -), 7.21–8.36 (m, 7H,
= 0.53 (toluene/acetone (5:3)).
.2.1.4. 4-(3-Bromopropoxy)-9H-xanthen-9-one was obtained as
white solid (yield 50%), mp 124–126 °C. Anal. Calcd for C16 Br:
(cm ): 1662,
6
, 300 MHz) d (ppm): 2.17–2.24 (m,
5.2.2.4. 4-(3N-(2-Amino-2-methyl-1,3-dihydroxypropyl)-2-hydroxy-
propoxy)-9H-xanthen-9-one hydrochloride (4) was obtained as white
2
2
2
2
2
ꢀ1
Ar–H). R
f
solid (yield 75%), mp 242–244 °C. IR (KBr)
1657, 1608, 1469, 1278, 1079, 751. R = 0.12 (methanol/ethyl acetate
(1:1)). Base of compound 4: Anal. Calcd for C20
v (cm ): 3410, 2921,
5
f
H
13
O
3
H
23NO
6
: C, 64.33; H,
ꢀ
1
1
C, 57.68; H, 3.93. Found: C, 57.73; H, 4.11. IR (KBr)
v
3
6.21; N, 3.75. Found: C, 64.18; H, 6.17; N, 3.47. H NMR (CDCl ,
1
1
606, 1492, 1468, 1327, 1270, 1228, 1062, 926, 749. H NMR
3
300 MHz) d (ppm): 0.87 (s, 3H, –CH ), 1.75 (s, 1H, –NH–), 2.61–2.77

1
350
H. Marona et al. / Bioorg. Med. Chem. 17 (2009) 1345–1352
O
OH
R1
O
1
) ClCH CH CH OH
2
2
2
K CO /acetone/EtOH
2
3
1
) epichlorhydrin/NaOH
2
) PBr /CHCl
3
3
O
O
O
O
O
R1
R1
O
O
Br
1
) aminolysis
K CO /toluen
1) aminolysis
n-propanol
2
3
2) HCl/EtOH
2) HCl/EtOH
O
O
R2
. HCl
R1
1
-8
Scheme 1. Synthesis of the tested compounds (1–8).
(
1
OH)
m, 1H, –NH–CH
2
-), 3.23 (t, J = 4.5 Hz, 4H, (–CH
H, –CH–OH), 4.08–4.21 (m, 2H, Ar–O–CH -), 4.33 (br s, 2H, (–CH
), 5.03 (d, J = 4.6 Hz, 1H, –CH–OH), 7.33–7.91 (m, 6H, Ar–H),
2
–OH)
2
), 3.90–3.92 (m,
f
(C-4a), 148.47 (C-4), 156.41 (C-4ba), 177.09 (C@O). R = 0.14
(methanol/ethyl acetate (1:1)).
5.2.2.6. 4-(3N-(2-Amino-2-methyl-1-hydroxypropyl)propoxy)-9H-
xanthen-9-one hydrochloride (6) was obtained as white solid (yield
2
2
–
2
8
.18 (d, J = 6.7 Hz, 1H, 8Ar–H).
.2.2.5. 4-(3-(Allylamino)propoxy)-9H-xanthen-9-one hydrochlo-
ride (5) was obtained as white solid (yield 72%), mp 185–187 °C.
Anal. Calcd for C19 Cl: C, 65.99; H, 5.83; N, 4.05. Found: C,
5
67%), mp 226–228 °C. Anal. Calcd. for C20
6.40; N, 3.71. Found: C, 63.51; H, 6.72; N, 3.79. IR (KBr)
3389, 2962, 2795, 1657, 1604, 1467, 1272, 1227, 1073, 753.
NMR (DMSO-d , 500.13 MHz) d (ppm): 0.96 (s, 6H, CH –CH–CH
1.94 (tt, J = 6.5 Hz, J = 6.7 Hz, 2H, –R–CH –R–), 2.71 (t, J = 6.7 Hz,
2H, –CH –NH–), 3.20 (s, 2H, –CH –OH), 3.36 (br s, 2H, –NH–, –
OH), 4.27 (t, J = 6.5 Hz, 2H, Ar–O–CH -), 7.38 (dd, J = 8.0 Hz,
4
H24NO Cl: C, 63.57; H,
ꢀ
1
1
v
(cm ):
H20NO
3
H
ꢀ1
6
1
d
3
5.98; H, 6.14; N, 3.82. IR (KBr)
656, 1606, 1469, 1343, 1277, 1075, 966, 756. H NMR (DMSO-
, 500.13 MHz) d (ppm): 2.25 (qu, J = 6.6 Hz, 2H, -R–CH -R-),
.18 (t, J = 6.5 Hz, 2H, –CH –NH–), 3.67 (ddd, J = 0.9 Hz, J = 1.2 Hz,
–CH@), 4.33 (t, J = 6.5 Hz, 2H, Ar–O–
–), 5.40 (ddt, J = 0.9 Hz, J = 1.5 Hz, J = 10.3 Hz, 1H, @CHH), 5.49
ddt, J = 1.2 Hz, J = 1.5 Hz, J = 17.2 Hz, 1H, @CHH), 5.96 (ddt,
v
(cm ): 3434, 2956, 2759,
6
3
3
),
1
2
6
2
2
2
2
2
J = 6.5 Hz, 2H, –NH–CH
CH
(
J = 6.6 Hz, J = 10.3 Hz, J = 17.2 Hz, 1H, –CH@), 7.41 (dd, J = 8.0 Hz,
J = 8.0 Hz, 1H, 2Ar–H), 7.52 (ddd, J = 1.1 Hz, J = 7.1 Hz, J = 8.0 Hz,
1
2
J = 8.0 Hz, 1H, 2Ar–H), 7.50 (ddd, J = 1.0 Hz, J = 7.1 Hz, J = 8.0 Hz,
1H, 7Ar–H), 7.52 (dd, J = 1.5 Hz, J = 8.0 Hz, 1H, 3Ar–H), 7.68 (ddd,
J = 0.5 Hz, J = 1.0 Hz, J = 8.5 Hz, 1H, 5Ar–H), 7.73 (dd, J = 1.5 Hz,
J = 8.0 Hz, 1H, 1Ar–H), 7.89 (ddd, J = 1.8 Hz, J = 7.1 Hz, J = 8.5 Hz,
1H, 6Ar–H), 8.20 (ddd, J = 0.6 Hz. J = 1.8 Hz, J = 8.0 Hz, 1H, 8Ar–H).
2
1
3
H, 7Ar–H), 7.56 (dd, J = 1.6 Hz, J = 8.0 Hz, 1H, 3Ar–H), 7.71 (ddd,
C NMR (DMSO-d
(R–CH –R), 38.11 (R–CH
CH –), 68.05 (–CH –OH), 116.43 (C-1), 117.65 (C-3), 118.31 (C-5),
6
, 500.13 MHz) d (ppm): 23.67 ((–CH
3 2
) , 30.12
J = 0.6 Hz, J = 1.1 Hz, J = 8.5 Hz, 1H, 5Ar–H), 7.77 (dd, J = 1.6 Hz,
J = 8.0 Hz, 1H, 1Ar–H), 7.92 (ddd, J = 1.8 Hz, J = 7.1 Hz, J = 8.5 Hz,
2
2
–NH–), 53.35 (–NH–C), 67.70 (Ar–O–
2
2
1
H, 6Ar–H), 8.21 (ddd, J = 0.6 Hz. J = 1.8 Hz, J = 8.0 Hz, 1H, 8Ar–H),
122.92 (C-8a), 121.98 (C-8b), 123.97 (C-2), 124.42 (C-7), 125.92
(C-8), 135.46 (C-6), 146.03 (C-4a), 147.83 (C-4), 155.34 (C-4ba),
176.02 (C@O). R = 0.06 (methanol/ethyl acetate (1:1)).
f
5.2.2.7. 5-(3N-(2-Amino-2-methyl-1-hydroxypropyl)-2-hydroxy-
propoxy)-3-chloro-9H-xanthen-9-one hydrochloride (7) was ob-
tained as white solid (yield 65%), mp 236–238 °C. Anal. Calcd. for
þ
13
9
.00 (br s, 2H, –NH2 –). C NMR (DMSO-d
-R), 44.71 (R–CH –NH–), 49.92 (–NH–CH
CH@), 67.58 (Ar–O–CH -), 118.02 (C-1), 118.98 (C-3), 119.50 (C-
), 122.02 (C-8a), 123.11 (C-8b), 123.59 (@CH ), 125.11 (C-2),
25.67 (C-7), 127.07 (C-8), 130.27 (@CH–), 136.66 (C-6), 147.13
6
, 500.13 MHz) d
(
ppm): 26.48 (R–CH
2
2
2
–
2
5
1
2

H. Marona et al. / Bioorg. Med. Chem. 17 (2009) 1345–1352
1351
C
20
H
23NO
5
Cl
2
: C, 56.08; H, 5.41; N, 3.27. Found: C, 56.05; H, 5.59;
in rats under anesthesia with thiopental (75 mg/kg, ip.) by iv. injec-
tion of adrenaline (20 g/kg). The studied compounds were admin-
ꢀ1
N, 3.20. IR (KBr)
v
(cm ): 3342, 1661, 1604, 1428, 1276, 1222,
075, 929, 754. H NMR (CDCl , 500.13 MHz) d (ppm): 1.29 (d,
J = 3.4 Hz, 6H, (–CH ), 3.05–3.15 (m, 1H, –CHH–NH–), 3.20–3.30
m, 1H, –CHH–NH–), 3.52 (d, J = 4.2 Hz, 2H, –CH –OH), 4.22–4.32
m, 2H, –CH –O–Ar), 4.32–4.39 (m, 1H, CH), 5.60 (t, J = 4.2 Hz,
H, –CH –OH), 5.95 (d, J = 4.7 Hz, 1H, –CH–OH), 7.42 (dd,
J = 8.0 Hz, J = 8.0 Hz, 1H, 2Ar–H), 7.55 (dd, J = 2.0 Hz, J = 8.5 Hz,
H, 7Ar–H), 7.61 (dd, J = 1.4 Hz, J = 8.0 Hz, 1H, 3Ar–H), 7.76 (dd,
J = 1.4 Hz, J = 8.6 Hz, 1H, 1Ar–H), 7.84 (d, J = 2.0 Hz, 1H, 5Ar–H),
l
1
1
3
istered via iv. route 15 min before adrenaline administration. The
criteria of antiarrhythmic activity were the lack of premature beats
and inhibition of cardiac arrhythmia in comparison to the control
group. The ED50 value was calculated according to the method of
3 2
)
(
(
1
2
2
2
1
2
Litchfield and Wilcoxon.
1
5.3.5. The effect on the arterial blood pressure
Male Wistar normotensive rats were anesthetized with thio-
pental (75 mg/kg) by intraperitoneally injection. The right carotid
artery was cannulated with polyethylene tub filled with heparin
in saline to facilitate pressure measurements using a Datamax
apparatus (Columbus Instruments, Columbus OH, USA). The stud-
ied compounds were injected into the caudal veins of rats after a
5 min stabilization period, at a constant volume of 1 mL/kg.
+
8
.20 (d, J = 8.5 Hz, 1H, 8Ar–H), 8.38 (t, J = 8.7 Hz, 1H, –NHH –),
+
13
8
.82 (t, J = 8.7 Hz, 1H, –NHH –). C NMR (DMSO-d
), 20.60 (–CH ), 44.28 (R–CH
–OH), 65.42 (–CH–OH), 71.41 (Ar–O–CH
17.27 (C-1), 118.25 (C-5), 118.85 (C-3), 119.94 (C-8a), 122.15
C-8b), 124.54 (C-2), 125.20 (C-7), 128.03 (C-8), 139.88 (C-6),
6
, 500.13 MHz)
–NH–), 59.85
–),
d (ppm): 20.41 (–CH
–NH–C), 64.68 (–CH
3
3
2
(
1
(
2
2
1
46.16 (C-4a), 147.52 (C-4), 155.69 (C-4ba), 175.46 (C@O).
= 0.06 (methanol/ethyl acetate (1:1)).
.2.2.8. 5-(3N-(2-Amino-2-methyl-1,3-dihydroxypropyl)-2-hydroxy-
R
f
5.3.6. Adrenoceptor radioligand-binding assay
5
The experiments were conducted on the rat cerebral cortex.
3
propoxy)-3-chloro-9H-xanthen-9-one hydrochloride (8) was obtained
as white solid (yield 70%), mp 218–220 °C. Anal. Calcd. for
[ H]prazosin (19.5 Ci/mmol,
1
a -adrenergic receptor) and
3
1
[ H]CGP-12177 (48 Ci/mmol, b -adrenergic receptor) were used.
C
20
H
23NO
6
Cl
2
: C, 54.06; H, 5.22; N, 3.15. Found: C, 54.06; H, 5.50;
The membrane preparation and the assay procedure were carried
out according to the published procedure with slight modifica-
tions. The brains of the rats were homogenized in 20 volumes of
ice-cold 50 mM Tris–HCl buffer (pH 7.6) and centrifuged at
20,000g for 20 min (0–4 °C). The cell pellet was resuspended in
Tris–HCl buffer and centrifuged again.
ꢀ1
22
N, 3.05. IR (KBr)
v
(cm ): 3352, 1599, 1426, 1276, 1071, 928, 756.
, 300 MHz) d (ppm): 1.20 (br s, 3H, –CH ), 3.15–3.21
m, 1H, –NH–CHH–), 3.36–3.40 (m, 1H, –NH–CHH–), 3.50–3.64 (m,
1
H NMR (CDCl
3
3
(
4
2
5
8
H, (–CH
H, (–CH
H, Ar–H), 8.09 (br s, 1H, –NHH –), 8.19 (d, J = 8.5 Hz, 1H, 8Ar–H),
2
–OH)
–OH)
2 2
), 4.19–4.32 (m, 3H, Ar–O–CH –CH(OH)), 5.47 (br s,
2
2
), 5.89 (d, J = 4.9 Hz, 1H, –CH–OH), 7.38–7.84 (m,
+
The final incubation mixture (final volume 300
240 lL membrane suspension, 30 l
l
L) consisted of
L of a [ H]prazosin (0.2 nM) or
H]CGP-12177 (0.2 nM) solution and 30 L buffer containing from
+
3
.62 (br s, 1H, –NHH –). R
f
= 0.11 (methanol/ethyl acetate (1:1)).
3
[
l
ꢀ11
ꢀ4
5
5
.3. Pharmacology
seven to eight concentrations (10 –10 M) of investigated com-
pounds. For measuring unspecific binding, phentolamine—10
l
M
3
.3.1. Animals and experimental conditions
(in the case of [ H]prazosin), and propranolol—1 lM (in the case
3
The studies were carried out on normotensive male Wistar rats
of [ H]CGP-12177), were applied. Radioactivity was measured in
a WALLAC 1409 DSA liquid scintillation counter (Perkin Elmer,
USA). All assays were done in duplicates. Radioligand binding data
were analyzed using iterative curve fitting routines with Graph-
weighing 170–350 g (Source: Animal House, Faculty of Pharmacy,
Jagiellonian University Medical College, Cracow, Poland, stocks
name: KRF: WI(WU). The animals were kept in plastic cages in a
room at a temperature of 20 ± 4 °C, under 12/12 h light/dark cycle
PAD/Prism, Version 3.0 (GraphPad ware, San Diego, CA, USA). K
i
(
light on from 7 a.m. to 7 p.m.). They were fed with granulated feed
values were calculated using the Cheng and Prusoff equation.23
(
standard laboratory pellets) and had free access to water. The con-
trol and study groups consisted of six animals each. All procedures
were conducted according to the Animal Care and Use Committee
guidelines, and approved by the Ethical Committee of Jagiellonian
University, Cracow.
5.3.7. Anticonvulsant and neurotoxicity assay
Initial evaluations for anticonvulsant activity were performed
due to the ADD (anticonvulsant drug development) program Epi-
lepsy Branch, National Institute of Neurological Disorders and
Stroke, National Institute of Health, Rockville, USA. The evaluations
of anticonvulsant activity included phase I test procedure. The
screens were performed in male Carworth Farms no. 1 (CF 1) mice
(18–25 g). In the phase I studies which deal with qualitative assay,
all the compounds were tested for activity in the MES test as well
as in the rotorod screen for TOX. The examined compounds were
suspended in 0.5% aq methylcellulose and then administered at
three dosage levels (30, 100 and 300 mg/kg) with anticonvulsant
activity observed 0.5 and 4 h after i.p. administration in mice.
The details of these procedures were published formerly.24
5
.3.2. Drugs
The tested compounds 1–8 were synthesized at the Department
of Chemical Technology and Biotechnology of Drugs UJ CM. Adrena-
line (Polfa, Warsaw, Poland), propranolol (Fluka Chemie AG, Seelze,
3
Germany), [ H]CGP-12177 (NEN-Du Pont, Warsaw, Poland),
3
[
H]prazosin (Amersham, Uppsala, Sweden), sodium heparin (Polfa,
Warsaw,Poland),thiopentalsodium(BiochemieGmbh,Vienna,Aus-
tria) were dissolved in saline. The tested compounds were adminis-
tered intravenously (iv.) at a constant volume of 1 mL/kg and were
investigatedat thewiderangeof dosesstartingat a doseof 10 mg/kg.
5
.4. Statistical analysis
5
.3.3. The effect on the normal electrocardiogram
Electrocardiographic investigations were carried out using AS-
The data were expressed as mean ± SEM. The statistical signifi-
PEL ASCARD B5 apparatus, (Aspel SA, Zabierzów, Poland) standard
lead II and paper speed of 50 mm/s. The ECG was recorded just
prior to and also 1, 5, and 15 min following the administration of
the compounds.
cance was calculated using one-way ANOVA. Differences were con-
sidered significant when p < 0.05.
Acknowledgements
5
.3.4. Adrenaline-induced arrhythmia
Prophylactic antiarrhythmic activity was determined according
The authors are grateful to Professor J. Stables for providing the
pharmacological data through the Antiepileptic Drug Development
program in National Institute of Health, Rockville, USA.
to the method of Szekeres and Papp.20 The arrhythmia was evoked

1
352
H. Marona et al. / Bioorg. Med. Chem. 17 (2009) 1345–1352
We are pleased to acknowledge the generous financial support
of this work by the Medical College of Jagiellonian University
Grant No. BBNK/ZDS/000717).
12. Fischer, W. Seizure 2002, 11, 285–302.
1
1
1
3. Marona, H. Pharmazie 1998, 53, 672–676.
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5. Marona, H.; P e˛ kala, E.; Antkiewicz-Michaluk, L.; Walczak, M.; Szneler, E. Bioorg.
Med. Chem. 2009, 17, 7234.
(
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